To study the mechanisms which regulate methionine biosynthesis, a search was made for inhibitors of specific steps in the pathway using Lemna as the experimental model. Nutritional uptake and labeling experiments were used to determine the site, extent and specificity of the metabolic block brought about by each inhibitor. A non-steady-state uptake experiment showed that molybdate interfered with the uptake of sulfate but provided no evidence of further impairment in organification of sulfate. Prevention of molybdate growth inhibition by cystine but not cystathionine or methionine provided support for our previous conclusion that transfer of sulfur from homcysteine to cysteine does not occur in plants. The metabolic block effected by lysine plus threonine was localized to the conversion of aspartate to homoserine. Propargylglycine produced growth inhibition due to inhibition of cystathionine gamma-synthase activity. Growth in aminoethoxyvinylglycine caused impaired cleavage of cystathionine to homocysteine. These results establish that each of these inhibitors brings about a methionine limitation, and thus validates their use in the study of regulation of the methionine pathway.